Veterinary Immunology.pptx

JIG-JIGA UNIVERSITY
COLLEGE OF VETERINARY MEDICINE
VETERINARY IMMUNOLOGY
Fikadu Alemu (DVM, MSc, Assist. Prof. of Veterinary Immunology)
College of Veterinary Medicine, Jig-jiga University

Chapter 1: The Basics of Immunology

Brief History of Immunology
• In Fifth century B.C. (B.C. 430)
• Thucydides in Athens first mentioned immunity to an infection that they called ‘plague’
• By 12th century
• Chinese observed that the individuals who recovered from small pox were resistant to further
attack.
• Then they deliberately infected infants by making small cut on the skin and rubbing the scabs
collected from the infected person.
• The children survived from the infection and they were protected from small pox later in their
life.
• Later with their experience they adopted a method of infecting children with the scabs collected
from the mildest cases of small pox (variolation) and the incidence of death due to small pox
dropped down from 20% to 1%.
• The method then spread to Europe and so on.
3

Documents show that as
early as 450 B.C,
the ancient Chinese
custom existed of having
children inhale powders
made from the
crusty skin lesions of
patients recovering
from smallpox

Cont…
• In 18th century
• Since the 9th Century, deaths was common among cattle due to Rinderpest (cattle plague) in
Western Europe
• (In 1754): soaking a piece of string with nasal discharge from the Rinderpest affected animal
and inserting into dewlap by making an incision in susceptible animals reduced the incidence
• In 1774,
• Benjamin Jesty, a farmer, inoculated his wife with vaccinia virus to protect her from small
pox
5

• In 1798,
• Edward Jenner (1749-1823, an English Physician) put the first step in the development of
immunology
• He observed that milk-maids who had recovered from cowpox never contracted smallpox
during their life
• inoculated a eight years old boy with blister fluid collected from cowpox and protected
him from severe small pox.
• This technique is called vaccination and used extensively to eradicate small pox from the
world.
• He is considered as the Father of Immunology.
6

African child with rash typical of smallpox on
face, chest, and arms. Smallpox, caused by the virus Variola major,
has a 30% mortality rate. Survivors are often left with disfi guring scars.
[Centers for Disease Control.]

Louis Pasteur (1822 – 1895)
• did extensive work on Fowl Cholera (Pasteurella multocida), Anthrax and Rabies and developed
vaccines against them.
• His work on development of vaccine against fowl cholera is accidental.
• He found out that some organisms, like Pasteurella multocida, loose their virulence when stored for
long time however they retain their ability to induce protection.
• Using the same principle, he developed vaccines for anthrax and rabies.
• He attenuated anthrax organisms by growing them at high temperature and rabies virus by drying
the affected rabbit brain.
• In 1885, administers the first vaccine to Joseph Meister
• He Established Pastuer institute at Paris, France
9

Pasteur’s Fowl Cholera Experiment: Birds inoculated with an aged culture of
P. multocida did not die. However, when subsequently inoculated with a fresh culture of
virulent P. multocida the birds were found to be protected 11

• Lious Pasteur (1822~1895).
• The genius of Pasteur carried
him to the solution of many
problems: the spoilage of beers
and wines, with the
accompanying pasteurization
process; the discovery of
anaerobic bacteria, virus
vaccines, and attenuation of
virulence; and studies of
spontaneous generation.
His studies in immunology
have rightly earned him
the position as father of
the science.

Von Behring
(1854~1917) discovered
the antitoxin and the
principles of antiserum
therapy. He established
one of the first
corporations to product
immunologic products.

• Robert Koch
(1843~1910)
• for his
investigations and
discoveries in
relation to
tuberculosis"

• Elie Metchnikoff
(1845~1916) converted his
discoveries of
phagocytosis into a
doctrine that gained many
disciples from his coterie
of students. He shared the
Nobel Prize with Ehrlich in
1908.

• Paul Ehrlich (1854~1915).
• Selective theories(Paul
Ehrlich,1900） The binding
like the fitting of a lock with
key,the side-chain
specificity was determined
before its exposure to Ag,
and the Ag selected the
appropriate side-chain
receptor.
• He shared the Nobel Prize
with Metchnikoff in 1908.

Clonal selection theory and immune tolerance

The clonal selection hypothesis

• Rodney R. Porter
(1917~1985) shared the
Nobel Prize in Physiology
and Medicine with
Edelman in 1972.
• Gerald M. Edelman
(1929~) was only 43
years of age when he
shared the Nobel Prize
with Porter in 1972.

Susumu Tonegawa is
a Japanese Scientist who
won the Nobel Prize for
physiology or medicine in
1987 "for his discovery of
the genetic principle for
generation of antibody
diversity"
Antibody Diversity

Peter C. Doherty Rolf M. Zinkernagel
”for their discoveries concerning the specificity of
the cell mediated immune defence”

The Nobel Prize in Physiology or Medicine 2011 was divided, one half
jointly to Bruce A. Beutler and Jules A. Hoffmann "for their discoveries
concerning the activation of innate immunity" and the other half to
Ralph M. Steinman "for his discovery of the dendritic cell and its role
in adaptive immunity".

A CHRONOLOGY OF IMPORTANT ACHIEVEMENTS IN IMMUNOLOGY
Year Scientists Involved Findings
1798 Edward Jenner Vaccination against small pox
1862 Ernst Haeckel Phagocytosis
1877 Paul Ehrlich Mast cells
1879 Louis Pasteur Attenuated vaccine for Fowl cholera,
1881 Louis Pasteur Attenuated vaccine for Anthrax and
swine erysipelas
1883 Eolie Metchnikoff Phagocytosis and cellular theory of
immunity
1885 Louis Pasteur Anti rabies vaccine
1888 Pierre Roux and
Alexander Yersin
Bacterial toxins
1890 Emil A von Behring
and Shibasaburo kitasato
Antitoxins in serum for diphtheria and
tetanus
1891 Robert Koch Delayed type of hypersensitivity
1894 Richard Pfeiffer and
Vasily Isaeff
Bacteriolysis
1894 Jules Bordet Complement and bacteriolysis
1900 Paul Ehrlich Antibody formation theory( side chain
theory)
1901 Karl Landsteiner ABO blood group
1901 Bordet and Gengou Complement Fixation Test
1902 Charles Richet
and Paul J. Portier
Anaphylaxis
1903 Nicholas Arthus Specific tissue destruction -Arthus
Phenomenon
1903 Sir Almoth Wright Opsonization (antibodies could help in
Phagocytosis)
1905 Von Pirquet Studied Serum Sickness
1906 Clemens Pirquet Allergy ( introduced the term allergy)
1912 Bacille, Albert Clamette and Camille
Guerin
B.C.G. vaccination against Tuberculosis
25

1921 Carl Prausnitz and Heinz Kustner Cutaneous allergic reaction
1930 Friedrich Breinl and Felix
Haurowitz
Template theory of antibody formation
1934 John Marrack Antigen-Antibody binding hypothesis
(Lattice theory)
1936 Grover Major Histocompatability Complex
(MHC)
1939 Tiselius and Kabat Antibodies are gamma globulins
1940 Karl Land Steiner and Alexander
Weiner
Identification of Rh antigen
1941 Albert H. Coons and others Fluorescence labeling
immunofluorescence
1942 Jules Freund and Katherine
McDermott
Adjuvants
1944 Peter Medwar and Frank
MacFarlane Burnet
Theory of acquired immunological
tolerance
1948 Orjan Ouchterlony and Stephen D
Elek
Double diffusion of antigen and antibody
in gels.
1948 Fagraeus Antibody production in Plasma B cells.
1952 James Riley and Geoffy Westt Histamine in Mast cell
1955-
59
Frank MacFarlane Burnet and Neils
K. Jerne
Clonal selection theory of antibody
formation
1955 Glick Bursa of Fabricius
1957 Isaacs and Lindenmann Discovery of interferon
1958 J. Dausset and F. Rapaport Histocompatibility antigens on human
leukocytes
1959 R.R. Porter, Gerald M.Edelman and
Alfred Nisonoff
Discovery of antibody structure
1961-
62
Miller and Good Discovery of thymus involvement in
cellular Immunity
1964-
68
Claman T and B cell co-operation in immune
response
1975 George Koehler and Caesar Milstein Monoclonal antibodies
1976 Susuma Tonegawa Gene arrangements in antibody
26

THEORIES OF IMMUNOLOGY
Cellular Immunity Theory
• In 1882 the Russian Zoologist Elie Metchnikoff (1845-1916) observed that when a rose thorn is
introduced into the larvae of a starfish, within a few hours it was surrounded by motile cells and
could be observed in the transparent starfish larvae.
• In 1883 he observed that fungal spores could be attacked by the blood cells in Daphnia, a
metazoan under microscope
• He extended his investigation to mammalian leukocytes, showing their ability to engulf micro-
organisms, a process which he termed phagocytosis
• This Put the basis for cellular immunity
27

Humoral Theory
• After Pasteur discovered that immunity can be produced by vaccination, it was soon recognized that
substances produced immunity are present in blood or in body fluid (Humor).
• Fodor in 1886 observed a direct action of immune serum on anthrax bacilli.
• George Nuttall in 1888 observed the bactericidal action of blood in certain animals.
• In 1889 Hans Buchnar showed that cells free serum is bactericidal and failed to have that effect after
heating at 55oC for one hour.
• The heat labile substance causing bactericidal effect was termed alexin (later named as cytolysin or
complement).
• In 1890 Von Behring and Kitasato demonstrated the neutralizing antitoxic activity by serum from
immunized animals with diphtheria or tetanus toxin. This was the first proof of humoral immunity.
28

Group discussion
•The Nature of Disease
•Types of Pathogenic Organisms
•Mechanisms of Disease by
Pathogens

• Pathogens, agents that cause disease, infect a wide
range of animals, including humans
• The immune system recognizes foreign bodies and
responds with the production of immune cells and
proteins
• All animals have innate immunity, a defense active
immediately upon infection
• Vertebrates also have adaptive immunity

The Nature of Disease
• Pathogenic Organisms
• Genetic Disorders
• Toxic Chemicals
• Other Environmental Factors
• Physical Damage to Organs
• Nutritional Disorders

Types of Pathogenic Organisms
• Viruses
• Bacteria
• Protozoan
• Fungi
• Parasites

Mechanisms of
Disease by Pathogens
• Utilization of host nutritional
resources
• Physical damage to host tissues
• Production of toxic substances
• Chromosomal and gene damage
• Body cells behave abnormally

• The term immunity, derived from the Latin “immunitas” (exempt), was adopted to
designate this naturally acquired protection against diseases such as measles or
smallpox.
• The emergence of immunology as a discipline was closely tied to the development of
microbiology.
• The impact of immunization against infectious diseases such as tetanus, pertussis,
diphtheria, and smallpox, are now either extinct or very rarely seen.
• Indeed, the impact of vaccination and sanitation on the welfare and life expectancy
of humans and animals has had no parallel in any other developments of medical
science

Cont…
• Immunology started to transcend its early boundaries and become a more
general biomedical discipline.
• Today, the study of immunological defense mechanisms is still an important
area of research,
• but immunologists are involved in a much wider array of problems, such as
self-nonself discrimination, control of cell and tissue differentiation,
transplantation, cancer immunotherapy, etc.
• The focus of interest has shifted toward the basic understanding of how the
immune system works in the hope that this insight will allow novel approaches
to its manipulation.

Cont…
• The living animal body contains all the components necessary to sustain life
• Because it is warm, moist and rich in many d/t nutrients
• As a result, animal tissues are extremely attractive to microorganisms that seek to invade the
body and exploit these resources for themselves.
• The magnitude of this microbial attack can be seen when an animal dies.
• The tissue Decomposes
• But the tissues of living, healthy animals are highly resistant to microbial invasion.
38

• This resistance is due to multiple interlinked defense mechanisms
• This defense is encompassed by the discipline of immunology and is the objective of this course.
• B/c effective resistance to infection is critical, the body do not rely on single defense
mechanisms.
• Multiple defense systems
• Effective against many d/t invaders
• Only destroy specific organisms
• Act at the body surface to exclude invaders
• Act deep within the body for those breached the outer surface
• Against bacteria, virus, fungus
• A failure in these defense will result in disease and possibly death
39
Cont…

Definitions
• Immunity means the body’s ability to resist infection or in other words the state of the body
which learns from the experience of past infection(s), how to deal more efficiently with
subsequent infections
• The immune system of an individual consists of its cells and molecules responsible for the
immunity
• Immune Response is the reaction of the body following an exposure to foreign antigen resulting in
the formation of antibodies and lymphokines
• Immunology is a branch of science that deals with the body’s resistance to infection or of altered
reactivity of the body following an exposure to a foreign substance or infection
40

Classification of the immune response

Response to Initial Infection and re-infection

Stages of Response to Infection

Course
of Typical
Acute
Infection

LYMPHOID SYSTEM
The immune system is organized into several special tissues which are collectively
termed as lymphoid or immune tissues.
• Those tissues that have evolved to a high degree of specificity of function are
termed as lymphoid organs.
1. Lymphoid cells- lymphocytes and plasma cells.
2. Lymphoid organs
- Primary (Central)
- Secondary (Peripheral).

Thymus
Present behind the upper part of the sternum.
Appearance- two lobes surrounded by a fibrous capsule.
Septa divides the glands to lobules with an outer cortex and
inner medulla.
Cortex- actively proliferating small lymphocytes.
Medulla- epithelial cells and mature lymphocytes in the
middle of which are Hassall’s corpuscles (whorl like
aggregates of epithelial cells).

BONE MARROW
• Lymphoid cells developing and maturing here are referred to as B cells (B
for Bursa of Fabricius or bonemarrow).
• Site for proliferation of stem cells and for the origin of pre-B cells and their
maturation to become immunoglobulin-producing lymphocytes.
• Like thymic selection during T-cell maturation, a selection process within
the bone marrow eliminates B cells with self reactive antibody receptor.

PERIPHERAL (SECONDARY) LYMPHOID ORGANS
Lymph nodes
Spleen
Mucosa associated lymphoid tissue

LYMPH NODES
 Placed along the course of lymphatics.
 Surrounded by a fibrous capsule from which trabeculae penetrates into the nodes.
 Outer cortex- accumulation of lymphocytes (primary lymphoid follicles) within which germinal centers
(secondary follicles) develop during antigenic stimulation.
 Follicle also contain dendritic macrophages.
 Inner medulla- lymphocytes, plasma cells and macrophages are arranged as elongated branching
bands (medullary cords).

LYMPH NODES
• Bursa dependent areas:
The cortical follicles and medullary cords that contain B-lymphocytes.
• Thymus dependent area:
Between the cortical follicles and medullary cords there is an ill-defined intermediate zone
(paracortical area) which contains T-lymphocytes.
• Functions:
. Filter for lymph, each group draining specific part of the body.
. Phagocytose foreign materials including microorganisms.
. Help in proliferation and circulation of T-cells and B-cells.
. They enlarge during local antigenic stimulation.

SPLEEN
Largest lymphoid organ.
Capsule from which trabeculae descends, dividing the organ into several interconnected
compartments.
• White pulp of spleen- constitute 3/4th of the organ.
Red pulp of spleen is the rest part.
Functions:
oFiltering and clearing of infectious organisms.
oServes as a ‘graveyard’ for affected blood cells.
oAs a reserve tank and settling bed for blood and as a systemic
filter for trapping circulating blood borne foreign particles.
oThe immunological function of spleen is primarily directed
against blood borne antigens.

EFFECTS OF SPLENECTOMY
• Depends on the age at which the spleen is removed.
• In children, splenectomy often leads to an increased incidence of bacterial
sepsis
• Splenectomy in adults has less adverse effects, although in some, it makes
the host more susceptible to blood-borne bacterial infection.

MUCOSA ASSOCIATED LYMPHOID TISSUE (MALT)
• The mucosa lining the alimentary, respiratory, genitourinary and other
lumina are endowed with a rich collection of lymphoid cell aggregates like
the Peyer’s patches or scattered isolated follicles- collectively called MALT.
- Gut associated lymphoid tissue (GALT): gut, adenoids, tonsils and colon.
- Bronchus associated lymphoid tissue (BALT): respiratory tract.

MUCOSA ASSOCIATED LYMPHOID TISSUE
(MALT)
• MALT contains lymphoid as well as phagocytic cells.
• Both B and T cells are present.
• While the predominant immunoglobulin produced in the mucosa is secretory
IgA, other immunoglobulin classes, lgG, 1gM and IgE are also formed locally.

CELLS OF THE IMMUNE SYSTEM
• Cells of the immune system are associated with the lymphatic system of the
body and its specialized cells.
• Lymphocytes of the lymphatic system are derived from stem cells of the bone
marrow.
• These undifferentiated precursor cells proliferate throughout life and replenish
the mature cells of the immune system.

LYMPHOCYTES
• Small, round cells found in peripheral blood, lymph, lymphoid organs and in
many other tissues.
• In peripheral blood it constitutes 20-45 per cent of the leucocyte population,
while in lymph and lymphoid organs they form the predominant cell type.
• Human body contains 1012 lymphocytes, approximately 109 of them being
renewed daily.
. Only about 1 per cent of the total body lymphocytes are present in the blood.

LYMPHOCYTES
CLASSIFICATION:
According to size:
1.SmaII (5-8 μm): most numerous, ‘hand-mirror’ form.
2.Medium (8-12 μm)
3.Large (12-15 μm)
• According to their life span:
• Short lived lymphocytes: about 2 weeks, they are the effector cells in the immune
response.
. Long lived lymphocytes: may last for 3 years or more, or even for life, these cells are
the storehouse for immunologic memory

LYMPHOCYTES
• Lymphocyte Recirculation
Policeman on beat patrol, ceaseless wandering of lymphocytes through the blood,
lymph, lymphatic organs and tissues.
Mount an immune response following antigenic introduction to any part of the
body or whenever necessary.
One cycle completed in about one or two days.
Recirculation lymphocytes are mainly T-cells.
B-cells tend to be more sessile.

LYMPHOCYTES
A number of surface antigens or markers have been identified on lymphocytes
and other leucocytes by means of monoclonal antibodies.
• These markers reflect the stage of differentiation and functional properties of
the cell.
• At the International Workshops for Leucocytes Differentiation Antigens order
was given by comparing the specificities of different antisera.
. When a cluster of monoclonal antibodies was found to react with a particular
antigen, it was defined as a separate marker and given a CD (Cluster
Differentiation) number.
. Over 150 CD markers have been identified so far.

T-CELLS
These are Thymus derived cells.
• Key players in adaptive immunity. Governs cell mediated immune response.
• 65-80% of the circulating pool of small lymphocytes.
Found in the inner subcortical regions but not in the germinal centers of the
lymph nodes.
• Longer life span (months or years) than B cells.
• On exposure to certain mitogens T cells can be stimulated to divide.

Development in the thymus
• All T cells originate from haematopoietic stem cells in the bone marrow.
• Haematopoietic progenitors derived from haematopoietic stem cells populate the thymus
and expand by cell division to generate a large population of immature thymocytes.
•, and are therefore classed as double-negative (CD4 CD8 )cells. The earliest thymocytes
express neither CD4 nor CD8As they progress through their development they become
double-positive thymocytes
. (CD4+ CD8+, and finally mature to single-positive (CD4+CD8- or CD4- CD8+) thymocytes
that are then released from the thymus to peripheral tissues.

Development in the thymus
• About 98% of thymocytes die during the development processes in the thymus by failing
either positive selection or negative selection, whereas the other 2% survive and leave the
thymus to become mature immunocompetent T cells.
• The thymus contributes more naïve T cells at younger ages.
• As the thymus shrinks by about 3% a year throughout middle age, there is a corresponding
fall in the thymic production of naïve T cells, leaving peripheral T cell expansion to play a
greater role in protecting older subjects.

HELPER (CD4 ) T CELLS
• About 65% of peripheral T cells
• Found mainly in the thymic medulla, tonsils and blood.
• Recognize a non peptide-binding portion of MHC Class I1 molecules.
• Hence CD4 T cells are restricted to the recognition of pMHC Class Il
complexes.

HELPER ( CD4 ) T CELLS
• Helper T cells become activated when they are presented with peptide antigens by
MHC class Il molecules that are expressed on the surface of Antigen Presenting Cells
(APCs).
• Once activated, they divide rapidly and secrete small proteins called cytokines that
regulate or assist in the active immune response.
. These cells can differentiate into one of several subtypes, including TH1, TH2, TH3,
TH17, or TFH which secrete different cytokines to facilitate a different type of immune
response.
. The mechanism by which T cells are directed into a particular subtype is poorly
understood, though signaling patterns from the APC are thought to play an important
role.

HELPER ( CD4 ) T CELLS
Involved in the induction and regulation of immune response.
They perform the following helper functions:
i. They help B cells to be transformed to plasma cells.
ii. CD8 T cells to become activated cytotoxic T cells, and
iii. Macrophages to mediate delayed type Hypersensitivity reactions.
. Main functions:
1. Help in the antigen specific activation of B cells and effector T cells.
2. Th-1 cytokines activates cytotoxic inflammatory and delayed hypersensitivity reactions.
3. Th-2 cells help in the production of interleukins which encourage production of antibodies
especially IgE.
4. Th-2 cytokines are associated with regulation of strong antibody and allergic response.

CYTOTOXIC (CD8 ) T CELLS
‘ Account for approximately one-third of all mature CD3 cells.
• Found mainly in the bone marrow and gut lymphoid tissue.
. Recognize a non peptide-binding portion of MHC Class I molecules (which is
present on the surface of nearly every cells of the body).
. Hence, CD8 T cells, also known as cytotoxic T cells are restricted to the
recognition of pMHC Class I complexes.

FUNCTIONS OF CD8 T-CELLS
• They kill: Virus-infected cells, Allograft cells, Tumor cells.
• T-cell mediated cytotoxicity is an apoptotic process that appears to be mediated
by two separate pathways:
i. Involving the release of proteins known as Performs, which insert themselves
into the target cell membranes forming channels.
These channels allow the diffusion of enzymes (granzymes) into the cytoplasm.
Granzyme induced apoptosis is calcium dependant.
ii. Signal delivery by cytotoxic cells to the target cells which require cell-to-cell
contact.

MEMORY T CELLS
i. Memory cells live for many years or have the capacity to
reproduce them.
ii. A large number of memory cells are produced, and so secondary response is
enhanced and ¡s greater than the primary response (‘memory against past
infection’).
iii.They are activated by small quantities of antigens and require less co-
stimulation than do the naïve and un activated T cells.
iv. Activated memory cells produce greater amounts of interleukins than do naïve
T cells when they are first activated.
Comprise two subtypes: Central memory T cells (TCM cells) and Effector memory
T cells (TEM cells).
i. Memory cells may be either CD4+ or CD8+.

REGULATORY T-CELLS
• Formerly known as suppressor T cells, are crucial for the maintenance of immunological tolerance.
Their major role is to shutdown T cell-mediated immunity toward the end of an immune
reaction and to suppress auto-reactive T cells that escaped the process of negative selection in the
thymus.
• Two major classes of CD4+ regulatory T cells have been described, including the naturally occurring
Treg cells and the adaptive Treg cells.
•.

REGULATORY T-CELLS
• Naturally occurring Treg cells arise in the thymus, whereas the adaptive
Treg cells may originate during a normal immune response
• Naturally occurring Treg cells can be distinguished from other T cells by
the presence of an intracellular molecule called FoxP3.
Mutations of the FOXP3 gene can prevent regulatory T cell development,
causing the fatal autoimmune disease

Innate Immunity:
The First Lines of Defense

A Joke by Teacher
•When I was teaching kindergarten
and had a cold, I would often get
laryngitis with it.
•One day a student asked me, “Do
you have a frog in your nose?”

• Microorganisms that are encountered daily in the life of a healthy
individual cause disease only occasionally.
• Most are detected and destroyed within minutes or hours by defense
mechanisms that do not rely on the clonal expansion of antigen-
specific lymphocytes.
• These are the mechanisms of innate immunity.

• Innate immunity relies on a limited number of receptors and secreted proteins
that are encoded in the germline and that recognize features common to many
pathogens.
• the innate immune system discriminates very effectively between host cells and
pathogens, providing initial defenses and also contributing to the induction of
adaptive immune responses.
• The importance of innate immunity is shown by the fact that defects in its
components, which are very rare, can lead to increased susceptibility to
infection, even in the presence of an intact adaptive immune system.

• The response to an encounter with a new pathogen occurs in three phases,
• When a pathogen succeeds in breaching one of the host's anatomic
barriers, some innate immune mechanisms start acting immediately.

The first defenses include:
several classes of preformed soluble molecules present in blood,
extracellular fluid, and epithelial secretions
Antimicrobial enzymes such as lysozyme begin to digest bacterial cell walls;
Antimicrobial peptides such as the defensins that lyse bacterial cell
membranes directly;
and a system of plasma proteins known as the complement system that
targets pathogens both for lysis and for phagocytosis by cells of the innate
immune system such as macrophages.

In the second phase of the response:
The innate immune cells sense the presence of a pathogen by recognizing -
pathogen-associated molecular patterns (PAMPs)-and become activated,
By themselves, neither the soluble nor the cellular components of innate
immunity generate long-term protective immunological memory.

The first lines of defense.
• Microorganisms that cause disease in humans and animals enter the body at
different sites and produce disease symptoms by a variety of mechanisms.
• Many different infectious agents can cause disease and damage to tissues, or
pathology, and are referred to as pathogenic microorganisms or pathogens.
• Once they have gained a hold, they require the concerted efforts of both innate
and adaptive immune responses to clear them from the body.
• In vertebrates, microbial invasion is initially countered by innate defenses that
preexist in all individuals and begin to act within minutes of encounter with the
infectious agent.

Epithelial surfaces of the body provide the first line of
defense against infection
• Our body surfaces are defended by epithelia, which impose a physical
barrier between the internal milieu and the external world that
contains pathogens
• Epithelia comprise the skin and the linings of the body's tubular
structures-the gastrointestinal, respiratory, and urogenital tracts.
• Epithelial cells are held together by tight junctions, which effectively
form a seal against the external environment.

• Infections occur only when a pathogen colonizes or crosses these barriers,
• The dry tough outer layer of the skin is a formidable barrier when not broken,
• Pathogen entry most often occurs through the vast area of epithelial surface
inside the body.
• The importance of epithelia in protection against infection is obvious when
the barrier is breached, as in wounds, burns,

• At this point there will be loss of the integrity of the body's internal
epithelia, in which cases infection is a major cause of mortality and
morbidity.
• In the absence of wounding or disruption, pathogens can set up an
infection by specifically adhering to and colonizing epithelial surfaces,
using the attachment to avoid being dislodged by the flow of air or fluid
across the surface.
• Some pathogens can also use surface molecules on the epithelial cells as
footholds to invade the cells or get into the underlying tissues.

Epidermis of skin
Bronchial ciliated epithelium
Gut epithelium

The complement system and innate immunity
• Complement is a collection of soluble proteins present in blood and other body
fluids.
• It was discovered in the 1890s by Jules Bordet as a heat-labile component of
normal plasma that augmented the opsonization and killing of bacteria by
antibodies, and so this activity was said to 'complement' the actions of antibodies.
• Opsonization refers to the coating of a pathogen by antibodies and/ or complement
proteins so that it is more readily taken up and destroyed by phagocytic cells.

• The complement system is composed of more than 30 different plasma
proteins, which are produced mainly by the liver.
• In the absence of infection, these proteins circulate in an inactive form.
• All complement proteins have final outcome of killing pathogen, either
directly or by facilitating its phagocytosis, and inducing inflammatory
responses that help to fight infection.

There are three pathways of complement activation.
classical pathway of complement activation.
alternative pathway, which can be activated by the presence of the
pathogen alone, and
lectin pathway, which is activated by lectin-type proteins that
recognize and bind to carbohydrates on pathogen surfaces.

• The three pathways of complement activation are initiated in different
ways.
• The lectin pathway is initiated by soluble carbohydrate-binding proteins-
mannose-binding lectin and the ficolins-that bind to particular carbohydrate
structures on microbial surfaces.
• Proteases associated with these recognition proteins then trigger the
cleavage of complement proteins and activation of the pathway.

• The classical pathway is initiated when the complement component C1,
which comprises a recognition protein (C1q) associated with proteases
(C1r and C1s), either recognizes a microbial surface directly or binds to
antibodies already bound to a pathogen.
• Finally, the alternative pathway can be initiated by spontaneous hydrolysis
and activation of the complement component C3, which can then bind
directly to microbial surfaces.

• These three pathways converge at the central and most important step in
complement activation.
• When any of the pathways interacts with a pathogen surface, an enzymatic
activity called a C3 convertase is generated.
• The C3 convertase is bound covalently to the pathogen surface, where it
cleaves C3 to generate large amounts of C3b, the main effector molecule of
the complement system, and C3a, a peptide that helps induce inflammation.

• Cleavage of C3 is the critical step in complement activation and leads directly or
indirectly to all the effector activities of the complement system.
• C3b binds covalently to the microbial surface and acts as an opsonin, enabling
phagocytes that carry receptors for complement to take up and destroy the C3b-coated
microbe.
• C3b can also bind to the C3 convertases formed by the classical and lectin pathways,
forming another multisubunit enzyme, a C5 convertase
• This cleaves C5, liberating the highly inflammatory peptide C5a and generating C5b.
• C5b initiates the 'late' events of complement activation, in which a further set of
complement proteins interact with C5b to form a membrane-attack complex on the
pathogen surface, creating a pore in the cell membrane that leads to cell lysis.

• The key feature of C3b is its ability to form a covalent bond with microbial
surfaces, which allows the innate recognition of microbes to be translated
into effector responses.
• Covalent bond formation is due to a highly reactive thioester bond that is
hidden inside the folded C3 protein and cannot react until C3 is cleaved.
• When C3 convertase cleaves C3 and releases the C3a fragment,
conformational changes occur in C3b that allow the thioester bond to react
with a hydroxyl or amino group on the nearby microbial surface
• If no bond is made, the thioester is rapidly hydrolyzed, inactivating C3b.

Distribution and function
of cell-surface receptors
for complement proteins.

Assembly of the membrane attack complex
generates a pore in the lipid bilayer membrane

The Induced Responses of Innate Immunity
• It is about the ancient system of pattern recognition receptors used by the
phagocytic cells of the innate immune system to identify pathogens and
distinguish them from self antigens.
• It also concerns how, the immediate destruction of pathogens, stimulation of
some of these receptors on macrophages and dendritic cells leads to their
becoming cells that can effectively present antigen to T lymphocytes, thus
initiating an adaptive immune response

Pattern recognition by cells of the innate immune system
• Regular patterns of molecular structure are present on many microorganisms but do not
occur on the body's own cells.
• Proteins that recognize these features occur as receptors on macrophages, neutrophils, and
dendritic cells, and as secreted molecules, such as the mannose-binding lectin (MBL)
• Pattern recognition receptors can be classified into four main groups on the basis of their
cellular localization and their function:
• free receptors in the serum (such as MBL),
• membrane-bound phagocytic receptors
• membrane-bound signaling receptors
• Cytoplasmic signaling receptors

Macrophages express receptors that
enable them to take up microbes by
phagocytosis.

Bactericidal
agents produced
or released by
phagocytes after
uptake of
microorganisms

Pathogen recognition and tissue damage initiate an inflammatory
response
• An important effect of the interaction between pathogens and tissue
macrophages is the activation of macrophages and other immune cells to
release
• small proteins called cytokines and chemokines ( chemoattractant cytokines), and
other chemical mediators that set up a state of inflammation in the tissue,
• attract monocytes and neutrophils to the infection
• An inflammatory response is usually initiated within hours of infection or
wounding.

• Macrophages are stimulated to secrete pro-inflammatory cytokines and
chemokines by interactions between microbes and microbial products and
specific receptors expressed by the macrophage.
• Inflammation has three essential roles in combating infection.
1. to deliver additional effector molecules and cells from the blood into sites of
infection, and so increase the destruction of invading microorganisms.
2. to induce local blood clotting, which provides a physical barrier to the
spread of the infection in the bloodstream.
3. to promote the repair of injured tissue.

Infection stimulates macrophages to release cytokines
and chemokines that initiate an inflammatory response

Monocytes circulating in the blood migrate into infected and
inflamed tissues

Toll-like receptors represent an ancient pathogen-recognition system
• Cytokine and chemokine production by macrophages is the result of stimulation of
signaling receptors on these cells by a wide variety of pathogen components.
• Of these receptors, the Toll-like receptors (TLRs) represent an evolutionarily ancient
host defense system.
• The receptor protein Toll was first identified as a gene controlling the correct dorso-
ventral patterning embryo of the fruitfly Drosophila melanogaster.
• But in 1996 it was discovered that in the adult insect, Toll signaling induces the
expression of several host –defense mechanisms, including antimicrobial
peptides such as drosomycin

Toll is required for
antifungal responses in
Drosophila melanogaster

The cellular locations of the mammalian Toll-like receptors

Killing by NK cells
depends on the
balance between
activating and
inhibitory signals

Chapter two and three
Humoral and Cell Mediated Immune Response

ANTIGEN
• Antigen is a substance which when introduced into the
tissues of a susceptible animal, it stimulates the formation
of specific neutralizing substances or antibody with which
it reacts specifically in some observable way or produced
lymphokines or both antibody and lymphokines
• Substances that can be recognized by the immunoglobulin
receptor of B cells, or by the T cell receptor when
complexed with MHC, are called antigens
161

Antigen Vs Immunogen
• The terms antigen and immunogen are often used
synonymously
• However, these terms antigen and immunogen, imply two
closely related entities
• Immunogen: is a molecule that provokes an immune
response
• Antigen: is a molecule which reacts with the antibody
produced or with the activated cellular constituents of
cell mediated immunity
163

Immunogenicity Vs Antigenicity
• Immunogenicity and antigenicity are related but distinct
immunologic properties that sometimes are confused
• Immunogenicity is the ability to induce a humoral
and/cell mediated immune response.
• Antigenicity is the ability of a molecule to be recognized by
an antibody or lymphocyte.
• All molecules possessing the property of immunogenicity also
possess antigenicity but the reverse is not always true.
• Molecules vary in their ability to act as antigens and stimulate
immune response.
164

Cardinal Features of an Immunogen
• Many different substances can induce immune responses.
The following characteristics influence the ability of a
substance to behave as an immunogen:
1. Foreignness
2. Molecular Size
3. Chemical Structure
4. Chemical complexity
5. Bio-degradability
165

1. Foreignness
• The cells that respond to antigens (antigen sensitive cells) are
selected so that their antigen receptors do not normally bind to
molecules originating within an animal (self antigens/ molecules)
• As a rule, only substances recognized as non-self will trigger the
immune response.
• The more the non-self it is, the better it is immunogenic.
• Microbial products and exogenous molecules are obviously non-self
and may be strongly immunogenic
166

2. Molecular Size
• The most potent immunogens are macromolecular
proteins [molecular weight (MW) of greater than 100,000
Daltons].
• Molecules smaller than 10,000 Daltons are often only
weakly immunogenic, unless coupled to an immunogenic
carrier protein.
• The larger the molecule, the more immunogenic it is likely
to be
167

The relative size of several significant antigens.
168
Size
Does
matter!

3. Chemical Structure
• Proteins and polysaccharides are among the most potent
immunogens, [although relatively small polypeptide chains,
nucleic acids, and even lipids can, if given the appropriate
circumstances, be immunogenic]
Proteins:
• Large heterologous proteins express a wide diversity of
antigenic determinants and are potent immunogens
• It must be noted that the immunogenicity of a protein is
strongly influenced by its chemical composition.
• basic proteins with clusters of positively charged amino
acids are strongly immunogenic (lysozyme and myoglobin) 169

Polypeptides:
• Hormones such as insulin and other polypeptides,
although relatively small in size (MW 1500), are usually
able to induce antibody formation when isolated from one
species and administered over long periods of time to an
individual of a different species
170

Polysaccharides:
• Polysaccharides are among the most important antigens
because of their abundant representation in nature.
• Pure polysaccharides, the sugar moieties of glycoproteins,
Lipopolysaccharides (LPS), glycolipid-protein complexes,
etc., are all immunogenic
Nucleic acids:
• Nucleic acids (RNA and DNA) usually are not immunogenic,
but they can induce antibody formation if coupled to a
protein to form a nucleoprotein.
171

4. Chemical complexity
• There appears to be a direct relationship between
antigenicity and chemical complexity:
• Chemically polymerized proteins are much stronger
immunogens than their soluble monomeric counterparts
• proteins immunogenicity increases with diversity of amino
acids
• Large complex molecules can be readily taken up by
macrophages
• Complex proteins are good immunogen than large
repeating polymers such as the lipids, carbohydrates and
nucleic acids.
Good antigens/ immunogens have complex structure
172

5. Bio-degradability
• All foreign materials are not capable of stimulating
immune response e.g. stainless steel pin, plastic heart
valves etc.
• The macromolecule which are degradable in nature can
act as immunogen.
• Plastic bags are inert organic polymers, not degradable
and they are not immunogen/antigen.
• The antigen molecule should be degraded and processed
to form suitable to trigger immune response.
173

Antigenicity
Dose
Route of
administration
Host
Genetics
Chemical
Complexity
Chemical
stability
Size
Foreignness
174
What makes a good antigen

FACTORS ASSOCIATED WITH THE INDUCTION OF AN IMMUNE
RESPONSE
• In addition to the chemical nature of the immunogen,
other factors strongly influence the development and
potency of an immune response
a) Genetic Background
b) Dose and Method of Antigen Administration
c) Use of Adjuvants
175

a) Genetic Background
• Different animal species and different strains of one given
species may show different degrees of responsiveness to a
given antigen.
• In humans, different individuals can behave as “high
responders” or “low responders” to a given antigen
• The genetic control of the immune response is confined
to the genes within the MHC
• The MHC gene products function to present processed
antigen to T cell thus playing a central role in determining
immunogenicity
176

b) Dose & Method of Antigen Administration
• An insufficient dose will not evoke an immune response, either
because it fails to activate enough lymphocytes or because it
induces a non-responsive state.
• An excessively high dose also can fail to induce a response
because it causes lymphocytes to enter a non responsive state
• The method of antigen administration has a profound effect on
the immune response
Route of administration:
• A given dose of antigen may elicit no detectable response when
injected intravenously, but a strong immune response is observed
if injected intradermally.
• The presence of dendritic cells in the dermis (Langerhans cells)
may be a critical factor determining the enhanced immune
responses when antigens are injected intradermally 177

Use of Adjuvants
• Adjuvants are substances that enhance the bodies
immune response (when administered along with
antigens)
• In contrast to carrier proteins, adjuvants are:
• often non-immunogenic and
• are never chemically coupled to the antigens
• Several factors seem to contribute to the enhancement of
immune responses by adjuvants, including:
• delayed release of antigen,
• nonspecific inflammatory effects, and
• the activation of monocytes and macrophages 178

Types of Antigens
• The antigens can be classified in two groups as:
• Microbial antigen and
• Non-microbial antigens
Microbial antigens
• It includes
• bacterial antigens,
• viral antigens and
• other microbial antigens
179

A. BACTERIAL ANTIGENS
There are two main groups:
1. Soluble antigens: they are substances produced by the bacteria,
which are excreted into the environment.
• E.g., enzymes, porins, heat shock proteins (HSP), exotoxins etc.
• Porins are proteins that form pores on the cell wall of gram
negative bacteria.
• HSP are produced in large quantities by bacteria undergoing
stresses.
• Exotoxins are highly immunogenic and stimulate the production
of Antitoxins
2. Cellular antigens: They are the structural units of bacterial cell.
Common bacterial antigens are the following:
i. Somatic (O) antigen:
 In gram negative bacteria (Salmonella, E. coli, Brucella etc.),
somatic antigens are composed of LPS–protein complex, which
are good antigen and produce good immune response.
 But O antigens are highly variable and thus immunity against
one ‘O” antigen will not confer immunity against bacteria
bearing other ‘O” antigens. 180

ii. Capsular (K) antigen: A variety of bacterial species have capsule
(e.g. Bacillus anthracis, E. coli, Salmonella spp. etc) which is
antigenic.
Capsule commonly consists of polysaccharides (e.g. K antigen of E.
coli) but some are composed of polypeptides (e.g. Poly-D-glutamic
acid in case of B. anthracis).
ii. Flagellar (H) antigen: Motile bacteria have flagella
(e.g. Salmonella spp., E. coli, Proteus spp.).
These Flagella are composed of protein (flagellin), which is antigenic.
ii. Fimbrial (F ) antigen: Fimbrial or Pili antigen are present on the
surface of bacteria
iii. Spore antigen: Bacterial spores.
(e.g. Bacillus spp., Clostridium spp. etc.,) especially the
exosporidium is antigenic.
181

B. VIRAL ANTIGENS
• Structural components (VP-Viral Protein) of the virus vary
in their size and complexity.
• Capsid protein and envelope (consists of lipoprotein and
glycoprotein) are antigenic.
• Examples, HN protein (glycoprotein) of Newcastle disease
virus.
183

C. Other Microbial Antigens
• Fungi, protozoa and worms  composed of different
types of antigens.
• They elicit immune response.
184

NON-MICROBIAL ANTIGEN
• The non microbial antigens can be classified as:
• cell surface antigens,
• auto antigens &
• miscellaneous
Cell surface antigens
• The surface of most of the cells is covered with different
antigens
• When these antigens are given to heterogeneous host an
immune response is mounted
• Eg. Blood group antigens, CD receptors of the leukocytes
185

Blood group antigens
The antigens found on the surface of red blood cells are called
blood group antigens or erythrocytes antigens.
• Most of the blood group antigens are either glycoprotein or
glycolipids, & they are the integral components of cell membrane
• They are not involved in antigen processing but they influence graft
rejection.
• There are several human blood group system e.g., ABO, MN, Rh,
Lewis, Kell, Duffy, Kidd, Colton etc.
• The ABO antigens in human are anion and glucose transporter
proteins.
• The M and C antigen of sheep RBC are associated with membrane
potassium pump and amino acid transport.
186

CD receptors of the leukocytes
• It refers to different molecules present on the surface of
lymphocytes, which perform specific function, and the
receptors can be identified by monoclonal antibodies
• Example CD8+ refers to T Cytotoxic cells.
CD4+ refers to T helper cells
187

Auto-antigen
• In certain circumstances, own body tissues develop
antigenic properties and antibody formed against the
antigen.
• The auto antigens are sequestrated without contact with
the lympho-reticular system but by any mishap when
antigens are released, they provide an opportunity to
produce autoantibody.
• Example: sperm, lens protein etc.
188

HAPTEN
• Haptens are usually non-protein substances of low
molecular weight having very little or no antigenic
property but acquire antigenicity when they are coupled
to a protein (carrier molecule).
• Haptens are incapable of inducing antibody formation by
themselves but can react specifically with antibodies.
• They are called partial antigen.
• It is possible to study the immune response of a well-
defined chemical by conjugating to a protein molecule.
189

• Hapten may be complex or simple molecule.
• Complex hapten is large molecular weight, polyvalent
compound.
• When they combine with antibody prepared against the
complete antigenic complex (new antigen) a visible
precipitation is formed.
• Simple hapten is a low molecular weight, univalent compound.
• When they combine with antibody prepared against complete
antigenic complex, no visible precipitation is formed.
• Examples: Penicillin, Dinitro Phenyl (DNP) etc.
191

INTRODUCTION
• An Antibody is a specific substance produced in the
body by B Cells in response to an antigen
• They are soluble proteins shed from B cell into
surrounding fluid following antigenic stimulation
• They bind specifically with antigen and try to destroy or
eliminate from the body
• Antibodies are found in many body fluids but more
concentration in serum
193

Nature of Antibodies
• Antibody molecules are glycoproteins
• Tiselius and Kabat (1938) subjected immune serum to
electrophoresis and separated its protein into four major
fractions - serum albumin, alpha (α), beta (β) and
gamma (γ) globulins.
• Most immunoglobulin (antibodies) are found in the
gamma fraction and some are in beta fraction
• Immunoglobulin are heterogeneous group of proteins
and constitute about 20% of plasma proteins
194

Structure of Antibodies
• Their structure is best illustrated by FOUR-CHAIN MODEL
• Antibody molecules are composed of two identical heavy chains
and two light chains.
– These chains are polypeptides made up of amino acids that are
arranged in a sequence.
• Disulphide bonds bind these chains.
– A single disulphide bond connects light chain with heavy chain
– two disulphide bonds connect the heavy chains.
• An antibody molecule has got two ends: an amino terminal and
caboxy terminal.
• The structure can be best explained as primary, secondary, tertiary
and quaternary structures.
195

196
Major surface receptors of B cells, their ligands and their function

1. Primary structure:
• It is the linear arrangement of amino acids in the polypeptides
chain.
• A heavy chain is composed of approximately 440 amino acids and
light chain has 220 amino acids.
• The sequence of amino acids from 1-107 in heavy and light chains
varies between antibody molecules and this region is called
variable (V) region.
• Whereas the amino acid sequence from 108- 440 in heavy chain and
from 108- 220 in light chain does not vary and this region is called
constant region (C).
• The arrangement of amino acid sequence in constant region also
follows a particular pattern of similarity. Based on the similarity in
arrangement the constant region is divided into domains. Each
domain has got approximately 100-110 amino acids.
199

Primary structure…..
• In an antibody molecule the variable region of heavy and light
chains have got only one domain named variable light (VL) and
variable heavy (VH).
• The constant region of light chain has got one domain and called
constant light (CL).
• But, the constant region of heavy chain has got 3 domains:
• The domains are indicated by number coupled with type of heavy
chains they have like: gamma, mu, alpha, delta or epsilon.
• Variability of the amino acid sequence in the V regions is not
random, but precisely organized.
• It is localized only in certain areas and these areas are referred as
hyper-variable regions or complementarity determining regions
(CDR)
200

• The four-chain model of structure is made clear by the
digestion of Ig molecules with enzymes: pepsin & papain
• Papain cleaves antibody molecule into three fragments
– Of them, two fragments can bind with antigens and are
referred as fragment antigen- binding (Fab).
– The remaining fragment can be crystallized and is referred
as fragment crystallizable (Fc).
– The Fc fragment does not have the property of binding
with antigen, & has got only antibody effector functions
• Pepsin provides an entirely different digestion pattern.
– Pepsin digestion provides a bivalent Fab and number of
small fragments of Fc region.
201

2. Secondary structure:
• it is the relationship between amino acids located some
distance apart in the primary structure
• Their interaction gives rise to periodic structural motifs
in protein molecule.
203

3. Tertiary structure:
• it describes the spatial relationship of amino acids that
are separated by great distances in the primary
sequence
• This refers to cysteine residues that form the intra chain
disulphide bridges
4. Quaternary structure:
• it reflects the interaction between distinct polypeptide
subunits of a multi-domain protein
• In an antibody molecule it reflects the biological activity
of antibody molecule.
204

CLASSES OF ANTIBODIES
• Generally there are five classes of antibodies
• Each class is distinguished by unique amino acid sequences in
the heavy-chain constant region, that confer class-specific
structural and functional properties.
• The major classes of antibodies are also known as isotypes.
• The antigenic determinants that determine the different
isotypes are called isotypic determinants.
• The isotypic determinants in the heavy chain are restricted to
constant region and termed in Greek letters as alpha, delta,
Epsilon, gamma and mu
205

• IgA – alpha (α) heavy chain
• IgM – mu (μ) heavy chain
• IgD – delta (δ) heavy chain
• IgE – epsilon (ε) heavy chain
• IgG – gamma (γ) heavy chain
206

• Antigenic determinants are also found on light chains,
and the light chains are classified into lambda (λ) and
kappa (κ) based on them.
• The antigenic determinants of light chains are restricted
to constant region
• The antigenic determinants of light chain are not useful
to identify major classes of antibodies since lambda and
kappa chains are found in all major types.
207

1. Immunoglobulin G (IgG)
• This is the major source of immunoglobulin secreted by plasma
cells found in the spleen, lymph nodes & bone marrow
• the most abundant class in serum, [constitute about 80% of the
total serum immunoglobulin]
• It has two heavy γ chains and two light chains of either κ or λ
types but not both
• is the smallest of all immunoglobulins and can easily escape the
blood vessels into the area of inflammation and distribution in
extra vascular & intravascular compartments.
• It is found in almost all tissue fluids & secretion except CSF
• It is the only immunoglobulin that can pass placental barriers
and found in newborn because of passive immunization.
208

• Based on antigenic and structural differences in heavy chains
the subclasses of IgG in humans are IgG1 (65-70%), IgG2 (23-
28%), IgG3 (4-7 %) and IgG4 (3-4%)
Functions
• Act as antitoxins
• Form antiviral antibodies
• Act as precipitins
• Bind to specific antigens such as those found on bacterial
surfaces.
• The presence of these antibody molecules on bacterial
surfaces can cause clamping (agglutination) and lead to
opsonization
• Act as complement fixing antibodies
• Provide passive immunity in new born animals or birds
• It is produced later to IgM, but provide long lasting immunity
209

Computer generated molecular model of IgG
211

2. Immunoglobulin M (IgM)
• IgM accounts for 5%–10% of the total serum immunoglobulin
with an average serum concentration of 1.5 mg/ml
• It occurs in the second highest concentration after IgG in
most mammalian serum
• Monomeric IgM, with a molecular weight of 180,000, is
expressed as membrane-bound antibody on B cells (as BCR).
• IgM is secreted by plasma cells as a pentamer in which five
monomer units are held together by disulfide bonds.
• The five monomer subunits are arranged with their Fc
regions in the center of the pentamer and the ten antigen-
binding sites on the periphery of the molecule.
212

• Each pentamer contains an additional Fc-linked polypeptide
called the J (joining) chain, which is disulfide-bonded to the
carboxyl-terminal cysteine residue of two of the ten chains
• IgM is the first immunoglobulin class produced in a primary
response to an antigen, and it is also the first
immunoglobulin to be synthesized by the neonate
Functions
• Biologically more active (effective) than IgG,
• A single molecule of IgM can cause immune hemolysis
whereas 1,000 IgG are required for the same effect
• IgM is 500-1000 times more effective than IgG in
opsonization, 100 times more effective in bactericidal action
and 20 times more effective in agglutination.
• It acts as complement fixing antibody.
214

3. Immunoglobulin A (IgA)
• IgA is the predominant immunoglobulin class in
external secretions such as breast milk, saliva, tears, and
mucus of the bronchial, genitourinary, and digestive
tracts.
• In serum, IgA exists primarily as a monomer, but
polymeric forms (dimers, trimers, and some tetramers)
are sometimes seen, all containing a J-chain polypeptide.
215

• IgA occurs in two forms: Serum IgA and Secretary IgA.
– Serum IgA is a monomer (with molecular weight 160 KDa).
– But secretary IgA (SIgA) usually found in mucosal surfaces
and in secretions is a dimer.
• It is formed by two IgA monomer joined at their carboxy
terminus of Fc region by “J chain” and also with a
secretary component (Sc).
• Secretary component is a glycoprotein (71 KDa),
synthesized in the epithelial cells of the mucous
membrane.
• It protects the IgA from digestion by intestinal proteases
216

Functions of IgA
• Provide local immunity to mucosal surface of respiratory
and intestinal tract against microbial invasion.
• It can prevent adherence of invading microbes to body
surfaces.
• It does not fix complement, but can activate alternative
complement pathway.
• It helps in phagocytosis and intracellular killing of
microorganisms
• It is a minor component in systemic humoral immunity but
plays a major role in mucosal immunity
• IgA antibodies found in gut contents or feces, thus are known
as copro antibodies. 218

5. Immunoglobulin E (Ig E)
• It is synthesized by plasma cells located beneath body surfaces
• It has two ε heavy chains and two light chains (either κ or λ).
• Constant region of heavy chain has four domains (CH 1 to CH4).
• Present in extremely low concentration in serum. Because of this,
it can’t act simply by binding and coating antigens, as the other
immunoglobins do.
• IgE antibodies mediate the immediate hypersensitivity reactions
that are responsible for the symptoms hay fever, asthma, hives,
and anaphylactic shock.
• It is largely responsible for immunity to parasitic worms.
• IgE has the shortest half life of all immunoglobulins (2 to 3 days)
and is readily destroyed by mild heat treatment.
219

Immunoglobulin D (IgD)
• IgD was first discovered when a patient developed a multiple
myeloma whose myeloma protein failed to react with anti-isotype
antisera against the then-known isotypes: IgA, IgM & IgG
• IgD has a serum concentration of 30 g/ml and constitutes about
0.2% of the total immunoglobulin in serum.
• IgD, together with IgM, is the major membrane bound
immunoglobulin expressed by mature B cells,
• Its role in the physiology of B cells is under investigation.
• No biological effector function has been identified for IgD.
• It has not been detected in all mammals (horses or rabbits
• IgD is a BCR mainly found attached to B cells)
221

Allotypes
• Individual animals show inherited variations in
immunoglobulin amino acid sequences.
• Thus the immunoglobulins of one individual may differ
from those of another individual of the same species.
• These inherited aa sequence variations in heavy chain
genes are called allotypes.
224

Idiotypes
• Structural variants found in immunoglubins results from
the variations in the amino acid sequences within the
variable domains of light and heavy chains. These
variants are called idiotopes.
• The collection of an immunoglobulin called its idiotype.
• some idiotypes may be located within the antigen
binding site, others are located on non-antigen binding
areas of the V domain.
225

Antigen-antibody interactions:
Are reversible specific non-covalent biochemical reactions:
– Hydrogen bonds (A chemical bond in which a hydrogen atom of one
molecule is attracted to an electronegative atom of another molecule)
– Electrostatic bonds(A valence bond in which two atoms, attracted by
electrostatic forces, transfer one or more electrons between atoms)
– Van der Waal forces (forces acting between nonbonded atoms or molecules)
– Hydrophobic bonds(The attractive force between molecules due to the close
positioning of non-hydrophilic portions of the two molecules
Can be represented by the formula:
K1=constant of association
K2=constant of dissociation
Ag + Ab Ag Ab
K1
K2

The affinity:
is the strength of the reaction between a single antigenic
determinant and a single combining site on the antibody
or it is the association constant for binding (KA)
KA= k1/k2
Valence: the number of epitopes
Avidity: is the collective affinity of multiple binding
sites(affinity+ Valence)

Types of Antigen-antibody reactions:
• Precipitation
• Agglutination
• Neutralization (Antitoxins)
• Opsonization
• Antibody-dependant cell-mediated
cytotoxicity
• The complement activation Membrane
attack complex

Consequences of Antibody Binding

PRECIPITATION
Is the reaction of soluble Ag with soluble Ab.
The reaction results in the formation of Ag-Ab
complexes (lattices)
Antigen
Antibody

The Quantitative Precipitation Reaction:
Varying amounts of Ag are mixed and incubated with
Constant volume of antisera
Precipitate is measured, amount of precipitate depends
on :
othe ratio of Ag : Ab
oThe Ab avidity
Plot in a curve, three zones are detected:
i. Zone of Ag excess : insufficient Ab  too small complexes
to precipitate
ii. Equivalence zone : large lattice is formed  visible
precipitates
iii. Zone of Ab excess : not enough Ag  too small complexes
to precipitate

AGGLUTINATION
Abs can bind and cross-link cells or particles 
aggregate formation
Entrap microbial invaders
IgM & IgA are the most suitable (IgG in sufficient amounts
can agglutinate cells)

Agglutination
RBC
RBC
RBC
RBC
IgM Antibody
IgG Antibody
RBC
RBC
RBC
RBC RBC
RBC
RBC

Applications of Agglutination
1. Agglutination/Hemagglutination:
a. Qualitative agglutination test
Determination of blood types or antibodies to blood group Ags
b. Quantitative agglutination test
Agglutination tests can also be used to measure the level of
antibodies to particulate antigens.(titration)
2. Passive hemagglutination: erythrocytes are coated
with a soluble antigen (e.g. viral antigen, a polysaccharide
or a hapten) and use the coated red blood cells in an
agglutination test for antibody to the soluble antigen

NEUTRALIZATION
Is the binding of Ab to microbial epitopes or soluble
molecules(e.g. toxins) which inhibits their binding to
host cells.
Abs are mostly IgG & IgA
 Used to identify toxins and viruses

OPSONIZATION
Is the process by which a pathogen is marked (tagged)
for ingestion and destruction by phagocytic cells

Antibody-dependant cell-mediated cytotoxicity
• Coating of an organism can attract phagocytic cells
as well as other cytolytic cells(NK cells, eosinophils)
• The organism may be: bacteria, protozoa, parasitic
worms
• These cells use cytolytic mechanisms to kill those
organisms

Destruction of Large Parasites by ADCC

Adaptive immunity
• Is specific for different microbial and non-microbial antigens
and increased by repeated exposures to antigen
• when it is mediated by B-lymphocytes it is called humoral
immunity and when mediated by T lymphocytes it is called
cell mediated immunity
• In contrast to innate immunity, when immunity develops as a
response to infection and adapts to the infection, it is called
adaptive immunity
• Has an extraordinary capacity to distinguish among different
closely related microorganisms or molecules and hence it is
called specific immunity and provides defense activity with
higher magnitude. 244

• The inflammatory process that is associated with many
infections triggers specific immune responses
• E.g. the production of a cytokine by antigen-ingested
macrophages that stimulate specific T lymphocyte against that
antigen
• The specific immunity increases the protective mechanisms of
innate immunity and makes it efficient in removing foreign
antigens.
• It can remember each antigen that has entered the body. This
immunologic memory helps in mounting a faster and efficient
removal when the same antigen enters subsequently.
• Specific immunity can also be called acquired immunity. Hosts
get this immunity as a result of exposure to an infection of
some intervention (vaccination).
245

Adaptive immunity can be acquired by two ways:
1. Active immunity
2. Passive immunity
Active immunity:
• it develops when the host’s body produced antibody in
response to foreign antigen.
• Active immunity develops slowly and persists for a long
time.
• It is highly protective
• Stimulates immunological memory
• Active immunity may be
Natural: in response to natural infection by infectious organisms
Artificial: This is produced by the host’s body in response to
inoculation of an antigen e.g., vaccination
246

Passive immunity:
• The antibody is prepared elsewhere and subsequently
introduced into host’s body.
• obtained either by transfer of serum or T cell
• It is established rapidly, but persists for short duration
• Provide moderate protection,
• there is no immunological memory
• The recipient of such transfer becomes immune immediately
• Passive immunity may be of two types:
Natural:
• Maternal antibody transfer to foetus (Transplacental)
• Colostrum antibody transfer through milk to neonates
Artificial
• By injection of immune serum in case of tetanus
• Transfer of lymphocyte or immune cells
247

The specific immunity can also be classified into:
humoral and cell mediated immunity
Humoral Immunity
is mediated by antigen specific blood glycoproteins called
antibodies
These antibodies are secreted by plasma cells that are
modified B-lymphocytes
it is very effective against antigens that are found in the
circulation
The antigen could be bacteria, exotoxin or virus
This immunity can be transferred to susceptible individual
by transferring the cell free portion of blood either plasma
or serum [passive immunity] 248

Cell mediated immunity
is mediated by antigen specific cells that are thymus derived cells
called T-lymphocytes.
There are two populations of T cells – T helper cells (TH cells) and
T cytotoxic cells (TC cells)
This immunity protects the host against intracellular antigens
like viruses, cancer antigens etc.
This immunity is also responsible for graft rejection in
transplants.
249

The differences between
humoral and cell mediated immunity
Humoral immunity Cell mediated
immunity
Antigen Extracellular
antigens
Intracellular
antigens
Responding
lymphocytes
B lymphocytes T lymphocytes
Effector mechanism Antibody mediated
elimination
Lysis of infected cell
Transferred by Serum T lymphocytes
250

FEATURES OF INNATE AND ADAPTIVE IMMUNITY
Characteristics Innate immunity Adaptive immunity
Specificity Common
structures shared by
groups of related
microbes and vital for
survival of organisms.
Recognize particular antigen
and develop specific immune
response.
Diversity Limited and germ line
encoded
Very large: large variety of
receptors is produced by
somatic recombination of
gene segments to recognize
antigens.
Memory No Yes
Non Reactivity
to self
Yes Yes
251

CARDINAL FEATURES OF ADAPTIVE IMMUNE RESPONSES
• The specific immune response comprises of humoral and
cell mediated responses.
• Both humoral and cell mediated responses have some
common fundamental properties
• Such common properties are referred as cardinal features
of immune responses and they are: specificity, diversity,
memory, specialization, self-limitaion and discrimination
between self and non-self
253

1. Specificity:
 Immune responses are specific for different structural components
of distinct antigens.
 The portions of antigen that are thus recognized by the immune
system (by individual lymphocytes) are called epitopes or antigenic
determinants
 An antigen may be composed of more than one epitope.
 Though antibody production is against all epitopes of an antigen,
maximum amount of antibodies are produced against only few
epitopes. Such epitopes are called immune-dominant epitopes.
 This specificity is due to the presence of membrane receptors on
the surface of lymphocytes that even recognizes small difference
between epitopes.
254

2. Diversity:
 The total number of antigenic specificities of the lymphocytes in an
individual is called lymphocyte repertoire
 Mammalian immune system can discriminate at least 109 distinct
antigenic determinants.
 Such diversity in a repertoire is due to variability in the structure of
antigen binding receptors or lymphocytes
255

3. Memory
when an individual is exposed to a foreign antigen the
event is recorded and the immune response against the
particular antigen is enhanced many folds during
subsequent exposures
This property of specific immunity is referred as
immunologic memory
The primary immune response to a particular antigen
takes 7-10 days to develop
Whereas the subsequent responses to the same antigen
occurs at a shorter period than 7-10 days and this is due to
immunologic memory.
256

Several features are responsible for immunologic memory.
oA particular antigen responsive retains its antigen receptors
during proliferation.
oTherefore each exposure the same antigen expands the clones of
lymphocytes specific for the antigen.
oA special group of cells are differentiated from antigen-stimulated
lymphocytes as a result of immune response.
oThese cells survive for a long time compared to other cells and
have mechanisms to overcome apoptosis, they maintain record
of antigen stimulation and are responsible for enhanced response
during subsequent exposure to the same antigen.
oThese specialized cells are called memory cells.
257

4. Specialization
 The immune system is adapted to respond in distinct and
special ways to different types of microbes like bacteria,
virus, etc.
 The immune mechanism followed for bacteria is different
from that for viruses.
 This specialization helps in eliminating microbial
infections.
258

5. Self-limitation
The immune response against a particular antigen is self-
limiting
The response wanes after some time
The antigen stimulated lymphocytes also perform their role for
a particular period
Once the antigen is removed they differentiate into memory
cells
However, numerous mechanisms for feedback regulations of
the immune responses are active during this quite phase
259

6. Discrimination of self from nonself (Tolerance):
 This is one of the most important aspects of immune system.
The individual’s immune system recognizes its own antigens
and does not mount an immune response.
 This immunologic unresponsiveness against individual’s own
antigen is referred as tolerance
 This tolerance is due to elimination of lymphocytes that react
for self-antigens or by functional inactivation of self-reacting
lymphocytes after their exposure to self-antigens
 When this self-tolerance fails antibody production occurs
against self-antigens (autologous antigens) that results in
production of autoimmune diseases
 Such immune response against self-antigens is referred as
autoimmunity
260

Phases of immune response
• The specific immune response that takes place after an
antigen stimulus can be divided into three phases:
1. Recognition phase
2. Activation phase and
3. Effector phase
1. Recognition phase:
This phase consists of binding of foreign antigens to
specific receptors on mature lymphocytes
This phase involves B-lymphocytes and T-lymphocytes
261

2. Activation phase:
the activation consists of sequence of events taking place in
induced lymphocytes due to specific antigen recognition
The two important changes taking place in lymphocytes due to
antigen recognition are proliferation and differentiation
As a result of proliferation, the specific clone of lymphocytes
increases in population amplifying the protective response
The B-lymphocytes differentiates into plasma cells and memory
cells.
Plasma cells have extensive ribosomes and produce large
quantities of antibodies.
The T-lymphocytes differentiates into different population with
each population having separate role in elimination of intra
cellular antigens.
262

3. Effector phase:
this phase is characterized by elimination of antigen against
which immune response was mounted
The lymphocytes that function in the effector phases are called
effector cells
Besides the lymphoid effector cells, the effector functions also
require non- lymphoid effector cells (like neutrophils,
eosinophils, etc.) and certain plasma proteins (complement)
The non- lymphoid effector cells effectively remove the
antigen-antibody complex formed
The plasma protein complement helps in lysis of the complex
Besides this, one of the products of complement cascade (C3b)
called opsonin forms a coat over the antigen and makes it more
vulnerable for phagocytosis (Opsonisation)
264

CELLULAR COMPONENTS OF ADAPTIVE IMMUNE SYSTEM
• The principal cells of the immune systems are lymphocytes, antigen
presenting cells (APCs) and effector cells
• Lymphocytes recognize foreign antigen and respond in two different
ways: Humoral & Cell mediated immunity
• B lymphocytes when they recognize extra cellular antigens, they
differentiated into antibody secreting cells and function as the
mediators of humoral immunity
• T- Lymphocytes recognize, intracellular antigen and destroy the
microbes or infected cells.
• They do not produce antibody. T lymphocytes do not respond to
soluble antigens but they recognize peptide antigen attached to host
proteins and produce different lymphokines to eliminate the
antigen.
• The third class of lymphocytes, natural killer (NK) cell is also involved
in innate immunity to remove intracellular organisms. 265

• For specific immune response, the antigen must be
captured and presented to specific lymphocytes.
• The cells, which perform this function, are called antigen-
presenting cells (APCs). They are mostly dendritic cells.
• Effector cells perform numerous functions to eliminate the
antigen.
• Activated T lymphocytes,
• mononuclear phagocytes and
• other leukocytes function as effector cells in different immune
responses.
266

Chapter 4
MHC AND DISEASE SUSCEPTIBILITY

MHC and Disease Susceptibility
• Some HLA alleles occur at a much higher frequency in those suffering from certain diseases than in
the general population.
• The diseases associated with particular MHC alleles include autoimmune disorders, certain viral
diseases, disorders of the complement system, some neurologic disorders, and several different
allergies.
• The association between HLA alleles and a given disease may be quantified by determining the
frequency of the HLA alleles expressed by individuals afflicted with the disease, then comparing these
data with the frequency of the same alleles in the general population.
• Such a comparison allows calculation of relative risk.
• A relative risk value of 1 means that the HLA allele is expressed with the same frequency in the patient
and general populations, indicating that the allele confers no increased risk for the disease.

Antigen Processing and Presentation
• Recognition of foreign antigen by a T cell requires that peptides derived from
the antigen be displayed within the cleft of an MHC molecule on the membrane
of a cell.
• Protein antigen degraded into peptides by a sequence of events called antigen
processing.
• The peptide- MHC complexes are transported to the membrane, where they are
displayed (antigen presentation).
• Class I MHC molecules bind peptides derived from endogenous antigens that
have been processed within the cytoplasm of the cell (e.g., normal cellular
proteins, tumor proteins, or viral and bacterial proteins produced within
infected cells).
• Class II MHC molecules bind peptides derived from exogenous antigens that are
internalized by phagocytosis or endocytosis and processed within the endocytic
pathway.

Cont…
• Processing of Antigen Is Required for Recognition by T Cells
• antigen recognition by B and T cells was basically similar.
• TH-cell activation by bacterial protein antigens was prevented by treating the
antigen presenting cells with paraformaldehyde prior to antigen exposure.
• However, if the antigen-presenting cells were first allowed to ingest the antigen
and were fixed with paraformaldehyde 1–3 h later, TH-cell activation still
occurred
• During that interval of 1–3 h, the antigen-presenting cells had processed the

Experimental demonstration that antigen
processing is necessary for TH-cell activation.
(a) When antigen-presenting
cells (APCs) are fixed before exposure to
antigen, they are unable to activate TH cells.
(b) In contrast, APCs fixed at least 1 h after
antigen exposure can activate TH cells.
(c) When APCs are fixed before antigen
exposure and incubated with peptide digests
of the antigen (rather than native antigen),
they also can activate TH cells.
TH-cell activation is determined by measuring
a specific TH-cell response (e.g., cytokine
secretion).

Separate antigen-presenting
pathways are utilized
for endogenous (green) and
exogenous (red) antigens.
The mode of antigen entry into
cells and the site of antigen
processing determine
whether antigenic peptides
associate with class I MHC
molecules in the rough
endoplasmic reticulum or with
class II molecules in endocytic
compartments.

Veterinary immunology Quiz 2
1. Mention at least three antigen – antibody reaction types.
2. Differentiate between humoral and cell mediated immune response.
3. State at least two characteristics of adaptive immune response.

Chapter 5. Immunological Methods (Diagnostic techniques)
Principles of Immunological Tests
• Immunoassay - the use of antibody to detect the presence of a specific antigen, or
vice versa
• In principle all immunoassays reflect the basic reaction between antibody and antigen
(analyte) as follows:
• Ab + Ag = Ab~Ag
• A fixed concentration of antibody reacts with varying concentrations of antigen
forming an antibody-antigen complex ('bound' antigen)
• Basis of immunoassays: Estimation of bound versus free components

Cont…
• Immunoassays/ Serological tests can be:
• Qualitative- positive or negative
• Semiquantitative
• Quantitative- quantification standards
• The reaction is reversible and will proceed to an equilibrium
• The proportions of this can be described by a constant, K
• As the value of K increases so too does the proportion of 'bound‘ complex

Reagents used in Serological Tests
• Serum
• Serum may be stored frozen and tested when convenient
• If necessary the serum can be depleted of complement activity – heating to 56oC
for 30 minutes
• Complement
• The normal constituent of all fresh serum
• The complement in fresh, unheated guinea pig serum is the most efficient in
hemolytic tests

cont…
• Serum used as a source of complement for serological applications should be stored frozen
in small volumes
• Once thawed, it should be used promptly; It should not be repeatedly frozen and thawed
• Antiglobulins
• Because immunoglobulins are complex proteins they re antigenic when injected into an
animal of a different species
• Antiglobulins are essential reagents in many immunological test
• Monoclonal Antibodies
• Are pure and specific

Assays Based on Immune (Ab-Ag) complex Formation
Ab-Ag interactions in solution
• Assays make use of the fact that large antigen–antibody complexes precipitate out of
solution, or appear as the visible clumping of bacteria or other cells, or activate
complement
• Cross-linking between Ag and Ab leads to clumping formation
• I. Cross-linking and the formation of immune complexes
• Because of bivalency, a single antibody molecule may use one antigen-combining site to
bind to its epitope on one molecule of antigen molecule, and the other antigen-combining
site to bind the identical epitope on a second antigen molecule

Cont…
• Each of these two antigen molecules may possess additional epitopes for additional
antibody binding, so that different antibody molecules mutually binding to this
antigen are said to be cross-linked
• Further cross connections between additional antigen and antibody molecules result
in the formation of an immune complex or lattice
• As more and more antigen and antibody molecules become cross-linked, they form
lattices large enough to precipitate out of solution and to become visible

Precipitation
• The classical precipitin reaction
• Ag solution (soluble Ag) + antiserum = Ag-Ab precipitates are formed
• Ag-Ag cross-linking= three dimensional lattice structure (Fc-Fc interaction)

Cont…
• Platform:
• Tubes - tube precipitation test
• Semisolid medium – immunodiffusion
• Enhanced precipitation – counter immunoelectrophoresis (in an electric
current)
• Speed and sensitivity of precipitation reaction

Immunodiffusion/ Agar gel diffusion
• Determinants of the reaction in agar medium are the relative concentrations of antigen
and antibody
• Single immunodiffusion:- Fixed antigen or antibody
• The other reactant is allowed to move and complex with it
• Double immunodiffusion:- both reactants are free to move toward each other and
precipitate
• Movement: linear or radial
• Application: quantitative & qualitative analysis of serum proteins
• When antigen and antibody diffuse toward each other within a semi-solid medium such as

Single radial immunodiffusion.
Relation of antigen concentration to size of
precipitation ring formed.
Antigen at the higher concentration [Ag1]
diffuses further from the well before it falls
to the level giving precipitation with
antibody near optimal proportions.

Radial immunodiffusion.
In this example, antiserum to IgA is
incorporated in agar and is used to measure
serum IgA levels.

Electroimmunodiffusion
• Coupling electrophoresis with diffusion
• One-dimensional double electroimmunodiffusion (counterimmunoelectrophoresis)
• - One-dimensional single electroimmunoduffusion (Laurell’s rocket electrophoresis)

Countercurrent immunoelectrophoresis
• Used as enhancement of precipitation
• Application: Antigens that migrate to the positive pole upon electrophoresis in agar
• Antigen and antiserum are placed in wells punched in the agar gel and a current
applied
• The antigen migrates steadily into the antibody zone forming a precipitin line

Agglutination
• Direct - to detect the pathogen (as Ag)
• An antibody is bound to the support
• Indirect - to detect an antibody against a pathogen
• An antigen is bound to the support
• Application: to identify bacteria, to type RBCs

Radioimmuno assay (RIA)
• The label is detected using a scintillation (gamma) counter
• The radioactive isotopes of inorganic molecules most commonly used as tags are:
• 125I and 131I (different radioisotopes of iodine), 57Co (cobalt), 75Se (selenium), and
32P (phosphorus)
• These radioisotopes can be covalently attached to protein (either antigen or
antibody) to generate tagged molecules of high specific activity that are easily
quantitated by specialized detection instruments

Fluorescent antibody tests/ Immunofluorescence
• Application:
• identification, quantitative enumeration, and sorting of a variety of cell types
• In some situations, the presence or precise localization of an antigen on an
individual cell is desired
• The signal visualization methods used with radioisotope and enzyme tags do not
provide the necessary degree of resolution
• However, certain dye molecules called fluorochromes can be visualized exactly in
the locations where they bind

Chapter 6. vaccination
• Vaccination is an effective strategy for restraining infections.
• Remarkably potent vaccines are those that are successful in provoking high-affinity
antibodies and memory cells.
• Most of the vaccines developed, function by inducing humoral immune response in
the host.
Attenuated and Inactivated Bacterial and viral vaccines
• Vaccines containing intact nonpathogenic microbes are made after attenuating the
virus or by killing the microbes taking care of their immunogenicity.
• Attenuated viral vaccines prove beneficial because they evoke effective adaptive
immune responses.

Cont…
• Live attenuated bacterial vaccines used nowadays offer protection but for small
duration while live attenuated viral vaccines elicit good response and long lasting
immunity.
• Viral vaccines perform better because of their adaption in cell culture.
• However live viral vaccines always face a potent risk of reversion to virulence and
hence safety is the main concern for such viruses.
• To minimize such risks inactivated vaccines are used such as influenza vaccine.

Purified Antigen (Subunit) Vaccines
• Subunit vaccines are those that contain a purified antigen and also needs to be given
along with an immunogenic enhancer called an adjuvant.
• Construction of subunit vaccine involves isolation of a specific protein from a virus
or bacteria before administration.
• Diphtheria and tetanus are the best examples of subunit vaccines.
Synthetic antigen vaccines
This concept involves identification of an epitope or an antigen and designing it in the
laboratory to be used in future as a vaccine.

Live viral vaccines involving recombinant viruses
• Recombinant viruses involve the insertion of genes that encode an antigen into a
noncytopathic virus, which provide immunity following its introduction to a
susceptible host.
• They offer protection by eliciting both innate and adaptive immune response.
However in some cases safety is the issue.
DNA vaccines
This founds the basis of most fundamental work being done in current times.
DNA vaccine strategy involves inoculation of a plasmid containing complementary
DNA encoding a protein antigen.
DNA vaccines elicit both humoral and cell mediated response even without any
adjuvant administration but their effectiveness needs more experimentation and
verification.

Vaccine development strategies

Chapter 7. Immune response to infectious diseases
•Pathogens & Disease
Pathogens are defined as microbes capable of causing host damage.
When host damage reaches a certain threshold, it can manifest
itself as a disease.
The evolution of an infectious disease in an individual involves complex
interactions between the pathogen and the host.

Important General Features of Immunity to Pathogens
• Microbial infections are best prevented by both innate and adaptive immune
responses.
• The innate immune system takes care of early defense while the adaptive
immune system offers a longer and potential response.
• As microbes differ a lot in their host attacking regime, their removal from the
affected patient requires efficient effector systems.
• The result of many microbial infections is decided by the balance between
microbial schemes for withstanding immunity and the host immune responses.
• Microbes have adapted several ways to combat the immune response.

Cont…
• Immunity against microbes performs almost similar to other defense
mechanisms.
• Although it is essential for host survival but sometimes it may cause damage to
the host tissue itself.
• Some microbes especially viruses have the potential to be latent.
• In such cases the host immune response does not allow the microbe to spread
but the microbes survive in the latent form, i.e. infection may prevail under
specific conditions like stress etc.

Immunity to bacteria
Immunity to extracellular bacteria
Extracellular bacteria are those that multiply and reside outside the host
cell.
These bacteria mainly affect the cells in two ways.
They either attack by causing inflammation and tissue damage or by
producing toxins.

Innate immunity to extracellular bacteria
• Innate immunity to extracellular bacteria essentially involves three processes.
Stimulation of phagocytes-
Phagocytes takes the help of surface receptors and Fc receptors to identify
extracellular bacteria and its opsonization respectively.
Most of these receptors are associated with promotion of phagocytic activity and
microbicidal activity.

Cont…
Induction of inflammatory response-
Antigen presenting cells like dendritic cells secrete cytokines which are
responsible for causing leukocyte infiltration at the site of inflammation.
Activation of complement system-
• Both gram positive and gram negative bacteria stimulate alternative pathway of
complement system and mannose expressing bacteria stimulate lectin pathway of
complement system by binding to mannose binding lectin.

Adaptive immunity to extracellular bacteria
• The immunity that plays major role against extracellular bacteria is the humoral
or antibody mediated immunity
• Usually polysaccharide antigens are prototypic thymus-independent antigens
and humoral immunity is the basic line of defense
• The antibodies in such cases defend the body by neutralization, opsonization,
phagocytosis and stimulation of complement system.
• Extracellular bacteria also stimulate the production of CD4+ helper T cells
which induces inflammation and phagocytic activity.

Immune evasion by extracellular bacteria
1) Polysaccharide antigens or encapsulated bacteria are more lethal as compared to a strain
devoid of capsule because they resist phagocytosis.
2) Capsulated bacteria inhibit alternate pathway of complement system due to the presence
of sialic acid.
3) One more way of evading immune response by extracellular bacteria is due to the genetic
edition of surface antigens.
E.g. surface antigen of some specific bacteria is contained in their pili.
Pili contain a protein antigen called “pilin” and this pilin undergoes gene variation.
Pili are the structures of bacteria responsible for bacterial adhesion to host cells.

Immunity to intracellular bacteria
• Some intracellular bacteria like pathogenic or facultative are able to multiply within
the phagocytes, so their elimination from the patients requires modified strategies.
Innate immunity to intracellular bacteria
Phagocytes and natural killer cells provide innate immunity to the intracellular
bacteria.
However some bacteria survive and multiply easily in the phagocytes, the phagocytes
need to be stimulated by the secretions of these bacteria in order to clear the
infection.

Cont…
The secretions from these bacteria are recognized by TLRs and cytoplasmic proteins
of the NOD-like receptor (NLR) family so that they stimulate the phagocytes to
degrade the invading bacteria.
In addition to the intracellular bacteria, activated natural killer cells produce IFN-γ,
which consecutively stimulates macrophages.
Although innate immunity provides protection from most of the bacteria but some
intracellular bacteria like Listeria monocytogenes need cell mediated immunity in
order to be eliminated from the body.

Adaptive immunity to intracellular bacteria
• T cell-mediated immunity plays a significant role in providing protection against
intracellular bacteria.
• CD4+ T-cells and CD8+ cytotoxic T lymphocytes are the two major forms of cell mediated
immunity that participate in phagocytosis or killing of infected cells, respectively.
• Both the, CD4+ T-cells and CD8+ cytotoxic T lymphocytes work together to provide
protection against the intracellular bacteria.
• Granulomatous inflammation acts as a marker for most of the infections due to
intracellular bacteria, which occurs because of T-cell and macrophage stimulation.

Dodging of immune system by intracellular bacteria
• Intracellular bacteria tend to dodge the immune system in many ways
comprising evading into the cytosol or preventing phagolysosome fusion and by
overpowering the reactive oxygen species by their microbicidal activity.
• These bacteria have the potential to cause chronic infections because they can
survive the phagocyte mediated elimination and thrive for years in the body and
may show reversion of the disease.

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